Although several members of protein disulfide isomerase (PDI) family support thrombosis, other PDI family members with the CXYC motif remain uninvestigated. ERp46 has 3 CGHC redox-active sites and a radically different molecular architecture than other PDIs. Expression of ERp46 on the platelet surface increased with thrombin stimulation. An anti-ERp46 antibody inhibited platelet aggregation, adenosine triphosphate (ATP) release, and αIIbβ3 activation. ERp46 protein potentiated αIIbβ3 activation, platelet aggregation, and ATP release, whereas inactive ERp46 inhibited these processes. ERp46 knockout mice had prolonged tail-bleeding times and decreased platelet accumulation in thrombosis models that was rescued by infusion of ERp46. ERp46-deficient platelets had decreased αIIbβ3 activation, platelet aggregation, ATP release, and P-selectin expression. The defects were reversed by wild-type ERp46 and partially reversed by ERp46 containing any of the 3 active sites. Platelet aggregation stimulated by an αIIbβ3-activating peptide was inhibited by the anti-ERp46 antibody and was decreased in ERp46-deficient platelets. ERp46 bound tightly to αIIbβ3 by surface plasmon resonance but poorly to platelets lacking αIIbβ3 and physically associated with αIIbβ3 upon platelet activation. ERp46 mediated clot retraction and platelet spreading. ERp46 more strongly reduced disulfide bonds in the β3 subunit than other PDIs and in contrast to PDI, generated thiols in β3 independently of fibrinogen. ERp46 cleaved the Cys473-Cys503 disulfide bond in β3, implicating a target for ERp46. Finally, ERp46-deficient platelets have decreased thiols in β3, implying that ERp46 cleaves disulfide bonds in platelets. In conclusion, ERp46 is critical for platelet function and thrombosis and facilitates αIIbβ3 activation by targeting disulfide bonds.

中文翻译：

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内质网蛋白 46 (ERp46) 在小鼠血小板功能和动脉血栓形成中的新作用。

尽管蛋白质二硫键异构酶 (PDI) 家族的几个成员支持血栓形成，但具有 CXYC 基序的其他 PDI 家族成员仍未得到研究。ERp46 具有 3 个 CGHC 氧化还原活性位点，其分子结构与其他 PDI 完全不同。血小板表面 ERp46 的表达随凝血酶刺激而增加。抗 ERp46 抗体抑制血小板聚集、三磷酸腺苷 (ATP) 释放和 αIIbβ3 激活。ERp46 蛋白增强 αIIbβ3 激活、血小板聚集和 ATP 释放，而失活的 ERp46 则抑制这些过程。在通过输注 ERp46 挽救的血栓形成模型中，ERp46 敲除小鼠的尾部出血时间延长，血小板积累减少。ERp46 缺陷的血小板降低了 αIIbβ3 激活、血小板聚集、ATP 释放和 P-选择素表达。这些缺陷可被野生型 ERp46 逆转，并被含有 3 个活性位点中任意一个的 ERp46 部分逆转。αIIbβ3激活肽刺激的血小板聚集被抗ERp46抗体抑制，并且在ERp46缺陷的血小板中减少。ERp46 通过表面等离子共振与 αIIbβ3 紧密结合，但与缺乏 αIIbβ3 的血小板结合不良，并在血小板激活时与 αIIbβ3 物理结合。ERp46 介导血块收缩和血小板扩散。ERp46 比其他 PDI 更强烈地还原 β3 亚基中的二硫键，并且与 PDI 不同，ERp46 在 β3 中独立于纤维蛋白原生成硫醇。ERp46 裂解 β3 中的 Cys473-Cys503 二硫键，暗示 ERp46 的靶标。最后，ERp46 缺陷的血小板 β3 中的硫醇减少，这意味着 ERp46 裂解血小板中的二硫键。总之，ERp46 对于血小板功能和血栓形成至关重要，并通过靶向二硫键促进 αIIbβ3 激活。